Cellulose, the major component of plant biomass, is considered the most abundant biopolymer. Bayer, E. A., Chanzy, H., Lamed, R., Shoham, Y. (1998) Cellulose, cellulases and cellulosomes. Curr. Opin. Struct. Biol. 8, 548-557. Certain microorganisms are able to convert the monomer of cellulose, glucose, into various products useful in the production of biofuels and other methods. Cellulose is highly stable, has a high storage potential, low cost, and plentiful supply. Based on these and other properties, cellulose and enzymes capable of degrading and hydrolyzing it are useful in the sequestration, storage, and production of bioenergy. Lynd L R, Weimer P J, van Zyl W H, Pretorius I S (2002), “Microbial cellulose utilization: fundamentals and biotechnology,” Microbiol Mol Biol Rev 66: 506-577.
Crystalline cellulose is composed of linear polymers of β1-4 linked glucose, held in a tightly crosslinked crystalline lattice by a high degree of intermolecular hydrogen bonding. This structure confers stability but also hinders efficient deconstruction of cellulose. Strategies for commercial depolymerization of cellulose typically combine pretreatment to disrupt the crystalline structure, followed by enzymatic hydrolysis. Hilden L, Johansson G (2004), “Recent developments on cellulases and carbohydrate-binding modules with cellulose affinity,” Biotechnol Lett, 26: 1683-1693. Disruption of the crystalline structure and chemical hydrolysis typically requires high temperatures and low pH. See Kim J S, Lee Y Y, Torget, R W. (2001) “Cellulose hydrolysis under extremely low sulfuric acid and high-temperature conditions, Appl. Biochem. Biotechnol. 91-93 331-340. Enzymatic hydrolysis generally occurs under milder conditions. The degree of pretreatment required and the expense of subsequent cleanup steps are affected by properties of the enzymes used.
Bacteria capable of degrading cellulose include those belonging to the genera Aquifex, Rhodothermus, Thermobifida, Anaerocellum, and Caldicellulosiruptor. A recombinant thermostable endoglucanase of Aquifex aeolicus produced in E. coli showed maximal activity at 80° C. and pH 7.0 with a half-life of 2 h at 100° C. (Kim J S, Lee Y Y, Torget, R W (2001). Cellulose hydrolysis under extremely low sulfuric acid and high-temperature conditions. Appl. Biochem. Biotechnol. 91-93. 331-340)). The endoglucanases produced by Anaerocellum thermophilum and Caldicellulosiruptor saccharolyticus are multidomain enzymes composed of two catalytic domains, linked to carbohydrate binding domains by proline-threonine-rich regions (Zverlov V, Mahr S, Riedel K, Bronnenmeier K (1998a), “Properties and gene structure of a bifunctional cellulolytic enzyme (CelA) from the extreme thermophile ‘Anaerocellum thermophilum’ with separate glycosyl hydrolase family 9 and 48 catalytic domains,” Microbiology 144 (Pt 2): 457-465; Te'o V S, Saul D J, Bergquist P L (1995), “celA, another gene coding for a multidomain cellulase from the extreme thermophile Caldocellum saccharolyticum,” Appl Microbiol Biotechnol 43: 291-296; Saul et al. 1990. The recombinant endoglucanase of Rhodothermus marinus has a pH optimum of 6.0-7.0 and a temperature optimum at 100° C. (Halldórsdóttir S, Thórólfsdóttir E T, Spilliaert R, Johansson M, Thorbjarnardóttir S H, Palsdottir A, Hreggvidsson G O, Kristjánsson J K, Holst O, Eggertsson G. (1998), “Cloning, sequencing and overexpression of a Rhodothermus marinus gene encoding a thermostable cellulase of glycosyl hydrolase family 12,” Appl Microbiol Biotechnol 49: 277-284). The aerobic thermophilic bacterium Thermus caldophilus also produces an endoglucanase which exhibits high activity on CMC with cellobiose and cellotriose as products (Kim D, Park B H, Jung B-W, Kim M-K, Hong S I, Lee, D S (2006) Identification and molecular modeling of a family 5 endocellulase from Thermus caldophilus GK24, a cellulolytic strain of Thermus thermophilus. Int J Mol Sci 7: 571-589). In contrast, high-temperature, crystalline deconstructing cellulases from hyperthermophilic Archaea are few in number, despite efforts to identify such enzymes. Hyperthermophilic enzymes that act on cellulose typically lack identifiable cellulose binding domains.
Thus there is a need for improved cellulases, including cellulases encoded by hyperthermophilic archaea, and cellulases having high stability and tolerance to a range of chemical and physical parameters, including cellulases with activity at high temperatures and over a broad range of temperatures and pH, cellulases with higher catalytic activity and rate of conversion, activity in the presence of salts, ionic detergents, sulfhydryl reagents, and ionic liquids. Provided are polypeptides, compositions and methods that meet this need.